Although arsenic is acknowledged to be highly carcinogenic to human skin, its action at the cellular and molecular level is not well understood. It is thought that arsenite-induced cell proliferation is one of the central events involved in its carcinogenic activity. Our preliminary data have revealed that treatment of cells with arsenite results in the activation of phosphatidylinositol 3-kinase (PI-3K) and downstream kinases, protein kinase B (Akt) and p70 S6 kinase (p70S6k), as well as an increase in cyclin D1 induction, cell proliferation and transformation in human keratinocytes. Considering the essential role of PI-3K and its downstream kinases in cyclin D1 induction, cell proliferation and transformation, the main hypothesis of this proposal is that PI-3K and its downstream signaling pathways play a pivotal role in arsenite-induced cyclin D1 expression, cell proliferation and transformation in human keratinocytes. The long-term goal is to elucidate the molecular mechanisms by which arsenite causes human skin cancer development. The overall aim of this proposal is to test the effect of arsenite on the PI-3K signal transduction pathway and its role in the regulation of cyclin D1 expression, cell proliferation and transformation in human keratinocytes. Especially, we will investigate this issue in accordance with the following testable hypotheses and specific aims (SA): SA 1: To test the hypothesis that PI-3K is required for the activation of Akt and p70S6k in arsenite-treated human keratinocytes. SA 2: To investigate whether PI- 3K/Akt/p70 s6k pathway plays a role in arsenite-induced cyclin D1 expression and cell proliferation in human keratinocytes. SA 3: To test the hypothesis that PI-3K/Akt/p70S6k pathway plays a role in arsenite-induced cell transformation in human keratinocytes. SA 4: To test the hypothesis that AP-1 and/or NFkappaB are downstream transcription factor(s) in human keratinocytes response to arsenite treatment. SA 5: To test the hypothesis that activation of AP-1 and/or NFkappaB is required for induction of cyclin D1 expression, cell proliferation and cell transformation by arsenite in HaCat cells. The significance of the research proposed in this application is that the results derived from the proposed studies will greatly facilitate the understanding of the molecular mechanism of cancer development caused by arsenite in human skin. A better understanding of signal transduction pathways leading to the induction of cyclin D1 expression, cell proliferation and transformation, will provide valuable information needed for designing more effective agents for the prevention and therapy of cancers caused by arsenite. Such agents could interfere with signaling pathways leading to cyclin D1 induction, cell proliferation and cell transformation.